Analyse d’un screening CRISPR dans des cellules tryptophane neg. Raphaële a fourni un fichier word détaillé qui explique la manip et les algorithmes utilisés.
Il y a trois fichiers MLE et trois RRA (triplicats biologiques) qui contiennent chacun un b score = une moyenne, un z score = un score qui prend en compte la moyenne et la déviation standard et des pval + wald tests + fdr.
Je vais pool les trois fichiers, avoir le score z des triplicats et déterminer les outliers puis les comparer à ceux que Raphaële a (pour contrôler que je fais pas du caca). Ensuite je prendrai les outliers recroisés entre le MLE et RRA pour pouvoir produire les différentes figures dont Raphaële a besoin.
files = list.files("./data/", pattern = "*mle_trp*")
for (i in 1:length(files)){
file = read_xlsx(paste0("./data/",files[i]))
file = file %>% filter(sgRNA >=4) %>%
dplyr::select(Gene, `treatment|beta`,`treatment|p-value`)
colnames(file) = c("Gene", paste0("mean_",i), paste0("pval_",i))
assign(paste0("mle_",i), file)
}
data = list(mle_1, mle_2, mle_3) %>%
purrr::reduce(full_join)
data_all =
data %>%
drop_na() %>%
rowwise() %>%
mutate(mean = mean(c(mean_1, mean_2, mean_3)),
sd = sd(c(mean_1, mean_2, mean_3)),
pval = mean(c(pval_1, pval_2, pval_3))) %>%
dplyr::select(Gene, mean, sd, pval)
# Replacing the pvalues = 0 into pval = 10^-5.5 to fit on the graph
data_all[data_all$pval==0,]$pval = 10^-5.5
data =
data_all %>%
filter(pval<= 0.05)
boxplot(data$mean)## [1] 0.4738267
outliers =
data %>%
filter(mean < quantile(data$mean)[2]|mean > quantile(data$mean)[4])
down =
data %>%
filter(mean < -0.3)
outliers %>%
head(15) %>%
knitr::kable()| Gene | mean | sd | pval |
|---|---|---|---|
| Mrpl28 | 1.1918667 | 0.1163599 | 0.0000032 |
| Ndufa1 | 1.1643667 | 0.0969317 | 0.0000181 |
| 37316 | 1.0765400 | 0.1310024 | 0.0000032 |
| Mrpl20 | 1.1647333 | 0.0715976 | 0.0000032 |
| Ndufa2 | 1.1362333 | 0.0476430 | 0.0000032 |
| Ptplb | 0.9026700 | 0.3572377 | 0.0004527 |
| Mrps25 | 1.1595667 | 0.1321792 | 0.0000032 |
| Agpat1 | 1.0188100 | 0.1424605 | 0.0000181 |
| Hsd17b10 | 0.9095200 | 0.1850416 | 0.0000362 |
| Ndufc2 | 1.1130333 | 0.0349172 | 0.0000032 |
| Gnb2l1 | 1.0756667 | 0.0651772 | 0.0000032 |
| Mrpl41 | 0.9315833 | 0.1756950 | 0.0000181 |
| Dbr1 | 0.9692067 | 0.1070501 | 0.0000181 |
| Tufm | 0.8323300 | 0.2199795 | 0.0000543 |
| Ncl | 0.6533667 | 0.3726756 | 0.0087642 |
## [1] 288 4
Quand je filtre pour pval < 0.05, j’ai 288 outliers pour les MLE compparé aux 261 de Raphaële. On va partir de la liste des outliers MLE+RRA qu’elle a fournie pour la suite.
Il faudrait retrouver les fichiers avec les pvaleurs, je peux pas plotter sinon.
| ID | Description | GeneRatio | BgRatio | pvalue | p.adjust |
|---|---|---|---|---|---|
| GO:0140053 | mitochondrial gene expression | 55/538 | 95/15811 | 0 | 0 |
| GO:0032543 | mitochondrial translation | 48/538 | 70/15811 | 0 | 0 |
| GO:0010257 | NADH dehydrogenase complex assembly | 35/538 | 47/15811 | 0 | 0 |
| GO:0032981 | mitochondrial respiratory chain complex I assembly | 35/538 | 47/15811 | 0 | 0 |
| GO:0033108 | mitochondrial respiratory chain complex assembly | 39/538 | 72/15811 | 0 | 0 |
| GO:0007005 | mitochondrion organization | 72/538 | 456/15811 | 0 | 0 |
| GO:0006399 | tRNA metabolic process | 42/538 | 155/15811 | 0 | 0 |
| GO:0009060 | aerobic respiration | 38/538 | 128/15811 | 0 | 0 |
| GO:0006119 | oxidative phosphorylation | 31/538 | 84/15811 | 0 | 0 |
| GO:0045333 | cellular respiration | 42/538 | 178/15811 | 0 | 0 |
| ONTOLOGY | ID | Description | GeneRatio | BgRatio | pvalue |
|---|---|---|---|---|---|
| BP | GO:0010257 | NADH dehydrogenase complex assembly | 28/170 | 35/538 | 0.00e+00 |
| BP | GO:0032981 | mitochondrial respiratory chain complex I assembly | 28/170 | 35/538 | 0.00e+00 |
| BP | GO:0033108 | mitochondrial respiratory chain complex assembly | 29/170 | 39/538 | 0.00e+00 |
| BP | GO:1901566 | organonitrogen compound biosynthetic process | 78/170 | 166/538 | 4.00e-07 |
| BP | GO:0006412 | translation | 59/170 | 116/538 | 8.00e-07 |
| BP | GO:0043043 | peptide biosynthetic process | 59/170 | 117/538 | 1.10e-06 |
| BP | GO:0043604 | amide biosynthetic process | 60/170 | 122/538 | 2.80e-06 |
| BP | GO:0006518 | peptide metabolic process | 59/170 | 120/538 | 3.70e-06 |
| BP | GO:0034645 | cellular macromolecule biosynthetic process | 62/170 | 130/538 | 7.20e-06 |
| BP | GO:0043603 | cellular amide metabolic process | 61/170 | 129/538 | 1.31e-05 |
| ONTOLOGY | ID | Description | GeneRatio | BgRatio | pvalue |
|---|---|---|---|---|---|
| BP | GO:0006357 | regulation of transcription by RNA polymerase II | 24/71 | 57/538 | 0e+00 |
| BP | GO:0065007 | biological regulation | 56/71 | 256/538 | 0e+00 |
| BP | GO:0006366 | transcription by RNA polymerase II | 25/71 | 63/538 | 0e+00 |
| BP | GO:0006355 | regulation of transcription, DNA-templated | 27/71 | 73/538 | 0e+00 |
| BP | GO:1903506 | regulation of nucleic acid-templated transcription | 27/71 | 73/538 | 0e+00 |
| BP | GO:2001141 | regulation of RNA biosynthetic process | 27/71 | 73/538 | 0e+00 |
| BP | GO:0051252 | regulation of RNA metabolic process | 30/71 | 88/538 | 0e+00 |
| BP | GO:0050789 | regulation of biological process | 53/71 | 238/538 | 0e+00 |
| BP | GO:0019219 | regulation of nucleobase-containing compound metabolic process | 32/71 | 103/538 | 1e-07 |
| BP | GO:0050794 | regulation of cellular process | 51/71 | 230/538 | 1e-07 |
| ID | Description | GeneRatio | BgRatio | pvalue | p.adjust |
|---|---|---|---|---|---|
| R-MMU-5389840 | Mitochondrial translation elongation | 73/369 | 83/8497 | 0.0e+00 | 0.0000000 |
| R-MMU-5368287 | Mitochondrial translation | 74/369 | 86/8497 | 0.0e+00 | 0.0000000 |
| R-MMU-5419276 | Mitochondrial translation termination | 73/369 | 85/8497 | 0.0e+00 | 0.0000000 |
| R-MMU-72766 | Translation | 88/369 | 220/8497 | 0.0e+00 | 0.0000000 |
| R-MMU-163200 | Respiratory electron transport, ATP synthesis by chemiosmotic coupling, and heat production by uncoupling proteins. | 53/369 | 116/8497 | 0.0e+00 | 0.0000000 |
| R-MMU-1428517 | The citric acid (TCA) cycle and respiratory electron transport | 60/369 | 163/8497 | 0.0e+00 | 0.0000000 |
| R-MMU-6799198 | Complex I biogenesis | 38/369 | 55/8497 | 0.0e+00 | 0.0000000 |
| R-MMU-611105 | Respiratory electron transport | 46/369 | 93/8497 | 0.0e+00 | 0.0000000 |
| R-MMU-189451 | Heme biosynthesis | 7/369 | 14/8497 | 7.0e-07 | 0.0000543 |
| R-MMU-163210 | Formation of ATP by chemiosmotic coupling | 7/369 | 17/8497 | 3.7e-06 | 0.0002245 |
| ID | Description | GeneRatio | BgRatio | pvalue | p.adjust |
|---|---|---|---|---|---|
| R-MMU-5419276 | Mitochondrial translation termination | 53/131 | 85/8497 | 0e+00 | 0.0e+00 |
| R-MMU-5368287 | Mitochondrial translation | 53/131 | 86/8497 | 0e+00 | 0.0e+00 |
| R-MMU-5389840 | Mitochondrial translation elongation | 52/131 | 83/8497 | 0e+00 | 0.0e+00 |
| R-MMU-72766 | Translation | 54/131 | 220/8497 | 0e+00 | 0.0e+00 |
| R-MMU-163200 | Respiratory electron transport, ATP synthesis by chemiosmotic coupling, and heat production by uncoupling proteins. | 38/131 | 116/8497 | 0e+00 | 0.0e+00 |
| R-MMU-1428517 | The citric acid (TCA) cycle and respiratory electron transport | 40/131 | 163/8497 | 0e+00 | 0.0e+00 |
| R-MMU-6799198 | Complex I biogenesis | 28/131 | 55/8497 | 0e+00 | 0.0e+00 |
| R-MMU-611105 | Respiratory electron transport | 32/131 | 93/8497 | 0e+00 | 0.0e+00 |
| R-MMU-163210 | Formation of ATP by chemiosmotic coupling | 6/131 | 17/8497 | 1e-07 | 2.2e-06 |
| R-MMU-8949613 | Cristae formation | 6/131 | 17/8497 | 1e-07 | 2.2e-06 |
| ID | Description | GeneRatio | BgRatio | pvalue | p.adjust |
|---|---|---|---|---|---|
| R-MMU-392518 | Signal amplification | 4/35 | 32/8497 | 0.0000080 | 0.0023261 |
| R-MMU-428930 | Thromboxane signalling through TP receptor | 3/35 | 23/8497 | 0.0001072 | 0.0118473 |
| R-MMU-418592 | ADP signalling through P2Y purinoceptor 1 | 3/35 | 24/8497 | 0.0001221 | 0.0118473 |
| R-MMU-418597 | G alpha (z) signalling events | 3/35 | 31/8497 | 0.0002659 | 0.0154777 |
| R-MMU-456926 | Thrombin signalling through proteinase activated receptors (PARs) | 3/35 | 31/8497 | 0.0002659 | 0.0154777 |
| R-MMU-8939211 | ESR-mediated signaling | 5/35 | 158/8497 | 0.0004315 | 0.0207295 |
| R-MMU-977444 | GABA B receptor activation | 3/35 | 40/8497 | 0.0005699 | 0.0207295 |
| R-MMU-991365 | Activation of GABAB receptors | 3/35 | 40/8497 | 0.0005699 | 0.0207295 |
| R-MMU-3371568 | Attenuation phase | 2/35 | 13/8497 | 0.0012496 | 0.0389705 |
| R-MMU-9006931 | Signaling by Nuclear Receptors | 5/35 | 208/8497 | 0.0014950 | 0.0389705 |
| ONTOLOGY | ID | Description | GeneRatio | BgRatio | pvalue |
|---|---|---|---|---|---|
| BP | GO:0010257 | NADH dehydrogenase complex assembly | 28/170 | 35/538 | 0.0000000 |
| BP | GO:0032981 | mitochondrial respiratory chain complex I assembly | 28/170 | 35/538 | 0.0000000 |
| BP | GO:0033108 | mitochondrial respiratory chain complex assembly | 29/170 | 39/538 | 0.0000000 |
| BP | GO:1901566 | organonitrogen compound biosynthetic process | 78/170 | 166/538 | 0.0000004 |
| BP | GO:0006412 | translation | 59/170 | 116/538 | 0.0000008 |
| BP | GO:0043043 | peptide biosynthetic process | 59/170 | 117/538 | 0.0000011 |
| BP | GO:0043604 | amide biosynthetic process | 60/170 | 122/538 | 0.0000028 |
| BP | GO:0006518 | peptide metabolic process | 59/170 | 120/538 | 0.0000037 |
| BP | GO:0034645 | cellular macromolecule biosynthetic process | 62/170 | 130/538 | 0.0000072 |
| BP | GO:0043603 | cellular amide metabolic process | 61/170 | 129/538 | 0.0000131 |
| BP | GO:0006119 | oxidative phosphorylation | 21/170 | 31/538 | 0.0000223 |
| BP | GO:0007005 | mitochondrion organization | 38/170 | 72/538 | 0.0000487 |
| BP | GO:0140053 | mitochondrial gene expression | 31/170 | 55/538 | 0.0000525 |
| BP | GO:0032543 | mitochondrial translation | 28/170 | 48/538 | 0.0000551 |
| BP | GO:0043933 | protein-containing complex organization | 48/170 | 99/538 | 0.0000752 |
| BP | GO:0045333 | cellular respiration | 25/170 | 42/538 | 0.0000932 |
| BP | GO:0009060 | aerobic respiration | 23/170 | 38/538 | 0.0001293 |
| BP | GO:0015980 | energy derivation by oxidation of organic compounds | 25/170 | 43/538 | 0.0001609 |
| BP | GO:0065003 | protein-containing complex assembly | 45/170 | 95/538 | 0.0002875 |
| BP | GO:0022900 | electron transport chain | 17/170 | 27/538 | 0.0005729 |
| BP | GO:0022904 | respiratory electron transport chain | 16/170 | 25/538 | 0.0006507 |
| BP | GO:0019646 | aerobic electron transport chain | 13/170 | 19/538 | 0.0008742 |
| BP | GO:0022607 | cellular component assembly | 52/170 | 120/538 | 0.0014491 |
| BP | GO:0046034 | ATP metabolic process | 21/170 | 38/538 | 0.0014779 |
| BP | GO:0042773 | ATP synthesis coupled electron transport | 14/170 | 22/538 | 0.0016035 |
| BP | GO:0042775 | mitochondrial ATP synthesis coupled electron transport | 14/170 | 22/538 | 0.0016035 |
| CC | GO:0005739 | mitochondrion | 138/171 | 253/541 | 0.0000000 |
| CC | GO:0098798 | mitochondrial protein-containing complex | 90/171 | 128/541 | 0.0000000 |
| CC | GO:0000313 | organellar ribosome | 53/171 | 72/541 | 0.0000000 |
| CC | GO:0005761 | mitochondrial ribosome | 53/171 | 72/541 | 0.0000000 |
Organize by theme
BP Mitochondrial gene expression (encompasses mitochondrial translation)
BP ATP metabolic process (encompasses proton-motive force driven ATP prod)
CC respiratory chain complex I
BP oxidative phosphorylation
| ID | Description | GeneRatio | BgRatio |
|---|---|---|---|
| R-MMU-5419276 | Mitochondrial translation termination | 53/132 | 85/8497 |
| R-MMU-5368287 | Mitochondrial translation | 53/132 | 86/8497 |
| R-MMU-5389840 | Mitochondrial translation elongation | 52/132 | 83/8497 |
| R-MMU-72766 | Translation | 54/132 | 220/8497 |
| R-MMU-163200 | Respiratory electron transport, ATP synthesis by chemiosmotic coupling, and heat production by uncoupling proteins. | 39/132 | 116/8497 |
| R-MMU-1428517 | The citric acid (TCA) cycle and respiratory electron transport | 41/132 | 163/8497 |
| R-MMU-6799198 | Complex I biogenesis | 29/132 | 55/8497 |
| R-MMU-611105 | Respiratory electron transport | 33/132 | 93/8497 |
| R-MMU-163210 | Formation of ATP by chemiosmotic coupling | 6/132 | 17/8497 |
| R-MMU-8949613 | Cristae formation | 6/132 | 17/8497 |
| R-MMU-189451 | Heme biosynthesis | 5/132 | 14/8497 |
| R-MMU-1592230 | Mitochondrial biogenesis | 6/132 | 27/8497 |
| R-MMU-189445 | Metabolism of porphyrins | 5/132 | 31/8497 |
| R-MMU-3371378 | Regulation by c-FLIP | 2/132 | 10/8497 |
| R-MMU-5218900 | CASP8 activity is inhibited | 2/132 | 10/8497 |
| R-MMU-69416 | Dimerization of procaspase-8 | 2/132 | 10/8497 |
| R-MMU-1362409 | Mitochondrial iron-sulfur cluster biogenesis | 2/132 | 11/8497 |
| R-MMU-202427 | Phosphorylation of CD3 and TCR zeta chains | 2/132 | 12/8497 |
| R-MMU-199220 | Vitamin B5 (pantothenate) metabolism | 2/132 | 13/8497 |
| R-MMU-389948 | PD-1 signaling | 2/132 | 13/8497 |
| R-MMU-140534 | Caspase activation via Death Receptors in the presence of ligand | 2/132 | 15/8497 |
| R-MMU-202433 | Generation of second messenger molecules | 2/132 | 19/8497 |
| R-MMU-5357769 | Caspase activation via extrinsic apoptotic signalling pathway | 2/132 | 19/8497 |
| R-MMU-71403 | Citric acid cycle (TCA cycle) | 2/132 | 21/8497 |
| R-MMU-5213460 | RIPK1-mediated regulated necrosis | 2/132 | 28/8497 |
| R-MMU-5675482 | Regulation of necroptotic cell death | 2/132 | 28/8497 |
| R-MMU-1852241 | Organelle biogenesis and maintenance | 6/132 | 212/8497 |
| R-MMU-9706019 | RHOBTB3 ATPase cycle | 1/132 | 10/8497 |
| R-MMU-264870 | Caspase-mediated cleavage of cytoskeletal proteins | 1/132 | 11/8497 |
| R-MMU-71406 | Pyruvate metabolism and Citric Acid (TCA) cycle | 2/132 | 47/8497 |
## R version 4.2.2 (2022-10-31 ucrt)
## Platform: x86_64-w64-mingw32/x64 (64-bit)
## Running under: Windows 10 x64 (build 19045)
##
## Matrix products: default
##
## locale:
## [1] LC_COLLATE=English_Belgium.utf8 LC_CTYPE=English_Belgium.utf8
## [3] LC_MONETARY=English_Belgium.utf8 LC_NUMERIC=C
## [5] LC_TIME=English_Belgium.utf8
##
## attached base packages:
## [1] stats4 stats graphics grDevices utils datasets methods
## [8] base
##
## other attached packages:
## [1] org.Mm.eg.db_3.16.0 AnnotationDbi_1.60.0 IRanges_2.32.0
## [4] S4Vectors_0.36.0 Biobase_2.58.0 BiocGenerics_0.44.0
## [7] ReactomePA_1.42.0 clusterProfiler_4.6.0 msigdbr_7.5.1
## [10] readxl_1.4.1 ggrepel_0.9.2 forcats_0.5.2
## [13] stringr_1.4.1 dplyr_1.0.10 purrr_0.3.5
## [16] readr_2.1.3 tidyr_1.2.1 tibble_3.1.8
## [19] ggplot2_3.4.0 tidyverse_1.3.2
##
## loaded via a namespace (and not attached):
## [1] shadowtext_0.1.2 backports_1.4.1 fastmatch_1.1-3
## [4] systemfonts_1.0.4 plyr_1.8.8 igraph_1.3.5
## [7] lazyeval_0.2.2 splines_4.2.2 BiocParallel_1.32.4
## [10] GenomeInfoDb_1.34.4 digest_0.6.30 yulab.utils_0.0.6
## [13] htmltools_0.5.3 GOSemSim_2.24.0 viridis_0.6.2
## [16] GO.db_3.16.0 fansi_1.0.3 magrittr_2.0.3
## [19] memoise_2.0.1 googlesheets4_1.0.1 tzdb_0.3.0
## [22] Biostrings_2.66.0 graphlayouts_0.8.4 modelr_0.1.10
## [25] svglite_2.1.1 timechange_0.1.1 enrichplot_1.18.3
## [28] colorspace_2.0-3 rappdirs_0.3.3 blob_1.2.3
## [31] rvest_1.0.3 textshaping_0.3.6 haven_2.5.2
## [34] xfun_0.34 crayon_1.5.2 RCurl_1.98-1.9
## [37] jsonlite_1.8.3 graph_1.76.0 scatterpie_0.1.8
## [40] ape_5.6-2 glue_1.6.2 polyclip_1.10-4
## [43] gtable_0.3.1 gargle_1.2.1 zlibbioc_1.44.0
## [46] XVector_0.38.0 graphite_1.44.0 scales_1.2.1
## [49] DOSE_3.24.2 DBI_1.1.3 Rcpp_1.0.9
## [52] viridisLite_0.4.1 gridGraphics_0.5-1 tidytree_0.4.2
## [55] reactome.db_1.82.0 bit_4.0.4 httr_1.4.4
## [58] fgsea_1.24.0 RColorBrewer_1.1-3 ellipsis_0.3.2
## [61] pkgconfig_2.0.3 farver_2.1.1 sass_0.4.2
## [64] dbplyr_2.2.1 utf8_1.2.2 labeling_0.4.2
## [67] ggplotify_0.1.0 tidyselect_1.2.0 rlang_1.0.6
## [70] reshape2_1.4.4 munsell_0.5.0 cellranger_1.1.0
## [73] tools_4.2.2 cachem_1.0.6 downloader_0.4
## [76] cli_3.4.1 generics_0.1.3 RSQLite_2.2.19
## [79] gson_0.0.9 broom_1.0.1 evaluate_0.18
## [82] fastmap_1.1.0 ragg_1.2.5 yaml_2.3.6
## [85] ggtree_3.6.2 babelgene_22.9 knitr_1.41
## [88] bit64_4.0.5 fs_1.5.2 tidygraph_1.2.2
## [91] KEGGREST_1.38.0 ggraph_2.1.0 nlme_3.1-160
## [94] aplot_0.1.9 xml2_1.3.3 compiler_4.2.2
## [97] rstudioapi_0.14 png_0.1-7 reprex_2.0.2
## [100] treeio_1.22.0 tweenr_2.0.2 bslib_0.4.1
## [103] stringi_1.7.8 highr_0.9 lattice_0.20-45
## [106] Matrix_1.5-1 vctrs_0.5.0 pillar_1.8.1
## [109] lifecycle_1.0.3 jquerylib_0.1.4 data.table_1.14.4
## [112] cowplot_1.1.1 bitops_1.0-7 patchwork_1.1.2
## [115] qvalue_2.30.0 R6_2.5.1 gridExtra_2.3
## [118] codetools_0.2-18 MASS_7.3-58.1 assertthat_0.2.1
## [121] withr_2.5.0 GenomeInfoDbData_1.2.9 parallel_4.2.2
## [124] hms_1.1.2 grid_4.2.2 ggfun_0.0.9
## [127] HDO.db_0.99.1 rmarkdown_2.18 googledrive_2.0.0
## [130] ggforce_0.4.1 lubridate_1.9.0